A biosensor is a sensor which utilizes a molecule recognizing ability of a biological material such as micro-organisms, enzymes, antibodies, DNA, RNA and the like, and applies a biological material as a molecule recognition element. To be specific, it utilizes a reaction which is caused when an immobilized biological material recognizes an objective substrate, such as enzyme consumption due to respiration of a micro-organism, enzyme reaction, light emission, and the like. Among various biosensors, an enzyme sensor has progressively come into practical use, and an enzyme sensor for glucose, lactic acid, uric acid, and amino acid, has been utilized in medical diagnostics or food industry.
For example, this enzyme sensor reduces an electron acceptor by an electron which is generated by a reaction of substrate included in a sample liquid as a specimen and enzyme or the like, and a measurement apparatus electrochemically measures a reduction quantity of the electron acceptor, thereby performing quantitative analysis of the specimen (for example, refer to the brochure of international publication No. 01/36953).
Hereinafter, a conventional biosensor will be described with reference to FIGS. 6 and 7. FIG. 6(a) is an exploded perspective view illustrating the structure of the biosensor, and FIG. 6(b) is a plan view of the biosensor shown in FIG. 6(a).
With reference to FIGS. 6(a) and 6(b), a biosensor 400 is composed of a first insulating support (hereinafter referred to as “first support”) 41 comprising polyethylene terephthalate or the like, a reagent layer 45, a spacer 46 having a cutout part 46a for forming a specimen supply path 47, and a second insulating support (hereinafter referred to as “second support”) 48 having an air hole 49. The spacer 46 and the reagent layer 45 are sandwiched between the first support 41 and the second support 48 to be integrally arranged.
A conductive layer 410 comprising an electrical conductive material such as a noble metal, e.g., gold or palladium, or carbon is formed on the surface of the first support 41 by employing a screen printing method, a sputtering evaporation method, or the like. The conductive layer 410 on the first support 41 is divided by plural slits, thereby providing a measurement electrode (hereinafter also referred to as “working electrode”) 42, a counter electrode 43, a detection electrode 44, and nearly arc shaped slits 414 and 415. Reference numerals 411, 412, and 413 are terminals of the measurement electrode 42, the counter electrode 43, and the detection electrode 44, respectively.
A reagent including enzyme, electron carrier, water-soluble polymer and the like, which uniquely reacts with a specific component in a sample liquid, is applied to the electrodes 42, 43, and 44, thereby forming the reagent layer 45, and spread of the reagent applied onto the electrodes 42, 43, and 44 is restricted by the nearly arc shaped slits 414 and 415 formed in the counter electrode 43.
The spacer 46 is further laminated thereon, and the specimen supply path 47 is formed by the rectangle cutout part 46a that is provided in the center of the front edge of the spacer 46.
The second support 48 is laminated and bonded onto the spacer 46 so that an end of the cutout part 46a of the spacer 46 leads to the air hole 49 provided in the second support 48.
Hereinafter, a description will be given of the procedure for measuring the content of the substrate in the sample liquid (specimen) using the conventional biosensor 400 constituted as described above.
Initially, a measurement apparatus (not shown) is connected to the biosensor 400, and a fixed voltage is applied to a space between the counter electrode 43 or the measurement electrode 42 and the detection electrode 44 by the measurement apparatus. Then, the sample liquid is supplied to the inlet of the specimen supply path 47 with the voltage being applied to the space between the two electrodes of the biosensor 400. The supplied sample liquid is drawn inside the specimen supply path 47 by capillary phenomenon, and passes on the counter electrode 43 and the measurement electrode 42 to reach the detection electrode 44. The sample liquid that has reached the detection electrode 44 dissolves the reagent layer 45. The measurement apparatus detects an electrical change that occurs between the counter electrode 43 or the measurement electrode 42 and the detection electrode 44, and starts a measurement operation.
In the above-described biosensor, at least two supports, i.e., the first and second supports 41 and 48 are bonded together to form the specimen supply path 47 through which the specimen is drawn into a space between the supports, and the reagent for analyzing the components of the drawn specimen is placed in the specimen supply path 47, and further, the air hole 49 that leads from the specimen supply path 47 to the outside is formed in at least one of the supports. When forming the air hole 49, in the conventional method, a portion of the second support is punched out by press working to open a hole having openings with flat peripheries as shown in FIGS. 7(a) and 7(b).
FIG. 7(a) is a perspective view illustrating the shape of the air hole that is produced by the conventional press working, and FIG. 7(b) is a cross-sectional view of the air hole.
In recent years, however, there has been a demand for a reduction in the amount of blood as a specimen to be supplied to the biosensor 400. With this demand, the size of the specimen supply path 47 is reduced, and consequently, the size of the air hole 49 formed in the second support 48 must be reduced.
However, reducing the size of the air hole 49 when it is formed by the press working causes a residue of the second support 48 that stays in the air hole 49 although a hole can be formed by pressing the second support 48. If a biosensor with the residue remaining in the air hole 49 is produced, a sufficient amount of specimen supplied cannot be drawn in the biosensor, resulting in a problem relating to measurement precision. The minimum diameter of the air hole which can be formed by the press working without the above-mentioned problem is 0.3 mm. Considering productivity, the diameter of the air hole by the current press working is 0.35 mm.
As a countermeasure against the above-mentioned problem, it is considered that a minute hole is formed not by pressing the second support 48 but by thermally melting the second support 48 with a laser or the like. According to this method, in contrast to the press working, no residue remains in the air hole 49, and further, a minute air hole can be formed.
However, when the size of the air hole to be formed in the second support 48 using the laser processing is too small, if the specimen supplied to the biosensor is a control solution or the like having a relatively low viscosity, since the speed at which the specimen is drawn into the specimen supply path 7 is too high as compared with the speed at which the air collected in the specimen supply path 47 gets out of the air hole 49, air bubbles remain in the specimen supply path 47 and thereby a sufficient amount of specimen cannot be drawn, resulting in a new problem that accurate measurement result cannot be obtained, although there occurs no problem when the specimen supplied to the biosensor is blood or the like having a relatively high viscosity.
Further, in the conventional biosensor 400 having the construction as described above, when a minute air hole 49 is formed and a specimen having a significantly low viscosity or a control solution is supplied to the air hole so as to be drawn into the specimen supply path 47, the supplied specimen as well as the dissolved reagent flow out of the air hole 49, whereby the measurement value to be detected by the measurement apparatus is lowered, resulting in a problem that highly accurate measurement value cannot be obtained.
In order to solve this problem, for example, it is considered that the surface of the second support 48 is coated with a water-shedding resin such as silicon to prevent the liquid specimen from flowing over the air hole 49.
However, prevention of overflow of the liquid specimen from the air hole 49 by the above-mentioned method takes much time and cost for applying the water-shedding material on the second support 48.